Ethylene induced cyanide resistant respiration in orchid petal cells

Respiration of isolated Aranda orchid petal cells increased markedly after cut flowers were treated with ethylene. An increase in respiration was observed 15 to 20h after treatment which was further enhanced in the presence of oxygen and ethylene. Ethylene induces the development of a cyanide resistant pathway in fully opened orchid flower tissues where the cyanide resistant capacity is negligible. However, there appears a shift back to the cyanide sensitive pathway some time after induction.     

Flavohemoglobin Hmp affords inducible protection for Escherichia coli respiration, catalyzed by cytochromes bo' or bd, from nitric oxide

Respiration of Escherichia coli catalyzed either by cytochrome bo' or bd is sensitive to micromolar extracellular NO; extensive, transient inhibition of respiration increases as dissolved oxygen tension in the medium decreases. At low oxygen concentrations (25-33 microm), the duration of inhibition of respiration by 9 microm NO is increased by mutation of either oxidase. Respiration of an hmp mutant defective in flavohemoglobin (Hmp) synthesis is extremely NO-sensitive (I(50) about 0.8 microm); conversely, cells pre-grown with sodium nitroprusside or overexpressing plasmid-borne hmp(+) are insensitive to 60 microm NO and have elevated levels of immunologically detectable Hmp. Purified Hmp consumes O(2) at a rate that is instantaneously and extensively (>10-fold) stimulated by NO due to NO oxygenase activity but, in the absence of NO, Hmp does not contribute measurably to cell oxygen consumption. Cyanide binds to Hmp (K(d) 3 microm). Concentrations of KCN (100 microm) that do not significantly inhibit cell respiration markedly suppress the protection of respiration from NO afforded by Hmp and abolish NO oxygenase activity of purified Hmp. The results demonstrate the role of Hmp in protecting respiration from NO stress and are discussed in relation to the energy metabolism of E. coli in natural O(2)-depleted environments.     

Influence of oxygen on thePseudomonas aeruginosa hydrogen cyanide synthase

Partially purified HCN Synthase (HCS) required exogenous electron acceptors for activity. Phenazine methosulfate (PMS) provided the greatest activity, whereas oxygen allowed only a limited response. TheP. aeruginosa secondary metabolite pyocyanin supported HCS-mediated cyanide production. HCN production by whole cells operated maximally at low oxygen levels, whereas moderate oxygen levels limited HCS activity. Respiration and cyanogenesis by whole cells were equally sensitive to azide; HCS was completely resistant.     

Respiratory activity of Acholeplasma laidlawii cells and its role in the active transport of carbohydrates]

The respiratory activity of the Acholeplasma laidlawii cells was studied in order to elucidate a possible mechanism of coupling of transport with energy. The respiration of the cells is stimulated by ethanol, glucose, NADH, lactate, and pyruvate. The substrates of the Krebs cycle have no effect on the respiration. The respiratory activity, stimulated by ethanol and glucose, is inhibited by the inhibitors of the respiratory chain, SH reagents, and the inhibitors of glycolysis. The results of experiments with inhibitors suggest that the respiratory chain in the A. laidlawii cells is reduced and terminated by flavoprotein. This is confirmed by the results of spectroscopic analysis of cytochromes. Respiration coupled with phosphorylation did not play any important role in the active transport of carbohydrates. Probably, the energy, necessary for the transport of carbohydrates, is supplied by the substrate phosphorylation. This explains the activation of respiration by glucose, which is so sensitive to arsenate. The respiration of the A. laidlawii cells is not stimulated by some carbohydrates (fructose, 3-O-methyl-D-glucose).     

Mitochondrial damage and inhibition of respiration in animal cell cultures treated with Triton WR-1339

Inhibition of cellular respiration by treatment with the nonionic detergent Triton WR-1339 was found to be related to the cytotoxic response of cell to the surfactant. Respiration of sensitive cell lines (AV-3 and HeLa) markedly inhibited by Triton concentrations as low as 125 μgm/ml. Conditionally sensitive lines (BHK-21 and L-929) were affected by 500 μgm/ml while the respiration of insensitive cultures (primary rat and chick embryo cells) was unaffected by this concentration. Macrocyclon, a cyclic analogue of Triton, failed to alter the respiration rate of any of the above cell cultures. The levels of isocitric and succinic dehydrogenases in sensitive and conditionally sensitive cells were depressed within 2 hours after treatment with 500 μgm/ml of Triton was initiated and by 6 hours the activity was only 25% of the untreated controls. Similar results were obtained with mitochondrial preparations from these cells. Enzyme levels in insensitive cells were unaffected by Triton treatment. Mitochondrial damage was the most striking characteristic noted in treated cells examined by electron microscopy. The mitochondria were quite distorted and had lost most of their cristae formation. This mitochondrial damage was seen in all cell types examined although the rate at which it occurred varied. With sensitive cultures, damage was pronounced within 6 hours after the addition of Triton while mitochondria from conditionally sensitive cells were not grossly affected until 48 hours and they appeared to repair the damage following the removal of Triton.     

Development of Respiration and Mitochondria in Mucor genevensis After Anaerobic Growth: Absence of Glucose Repression

Respiration and mitochondria in Mucor genevensis, a facultatively anaerobic dimorphic mold, have been studied in aerobically and anaerobically grown cells and in anaerobically grown cells adapting to aerobic conditions. Respiration in hyphae continues at a high level during aerobic growth but drops rapidly on exhaustion of glucose. In anaerobically grown yeastlike cells, containing no recognizable aerobic cytochromes, a small cyanide-insensitive respiration occurs. Mitochondria with well defined cristae are visible in negative contrast after KMnO(4) fixation of stringently anaerobic cells containing low amounts of fatty acid of which 10% or less are unsaturated. On aeration of anaerobically grown cells, respiratory capacity and cytochromes develop rapidly, even in the presence of 10% glucose, indicating that glucose does not repress development of respiration. However, mycelium formation by adapting yeastlike cells is repressed by high glucose concentration. In adapting cells, apparent changes in mitochondrial ultrastructure appear to be more related to changes in fixation properties of cells than to changes in the structure of mitochondria.     

Changes in cell morphology of Listeria monocytogenes and Shewanella putrefaciens resulting from the action of protamine.

Protamine, which is an antibacterial basic peptide, was shown to alter the cell morphology of Listeria monocytogenes and Shewanella putrefaciens. Atomic force microscopy revealed that protamine smoothed the surface of cells, formed holes in the cell envelope, and caused fusion of S. putrefaciens cells. Immunoelectron microscopy of protamine-treated cells of both L. monocytogenes and S. putrefaciens showed great damage to the cell wall and condensation of the cytoplasm. Respiration of the cells was decreased due to treatment with sublethal concentrations of protamine, probably due to leakage or loss of cell envelope potential. It was concluded that protamine disrupted the outer surface structure and condensed the cytoplasm of sensitive cells and, in sublethal concentrations, altered membrane structures, thereby eliminating respiration.     

The effects of triphenyltin chloride on respiration and photosynthesis in the green algae Enteromorpha intestinalis and Ulothrix pseudoflacca

Abstract Respiration and photosynthesis in zoospores of Enteromorpha intestinalis (L.) Link and Ulothrix pseudoflacca Wille showed greater sensitivity to inhibition by triphenyltin chloride than in vegetative tissue. Photosynthesis in zoospores and vegetative tissue of U. pseudoflacca was more resistant to triphenyltin chloride inhibition than in zoospores and vegetative tissue of E. intestinalis. Respiration in zoospores and vegetative tissue of both species was equally sensitive to triphenyltin chloride.     

Radiation Damage to Bull Sperm Motility. II: Proton Irradiation and Respiration Measurements

Diluted bull semen samples were bombarded with a 24 Mev proton beam. Dose response curves for the fraction of cells which survived the bombardment and for the average velocity of the surviving cells were measured. Target theory indicated a cross section of the sensitive volume of 2.1 × 10^-10 cm². Respiration measurements showed that the oxidative phosphorylation in the sperm remained coupled after the bombardments. The efficiency with which free energy from ATP hydrolysis was converted into mechanical work by the sperm was found to decrease after proton bombardment. The half-value dose for this effect was two and a half times higher than the half-value dose for motility damage. These respiration measurements indicate that the damage due to the bombardment is not to the metabolic system or to the contractile system in the sperm flagellum, but to a control system for the motility. The results of the target theory shows that this control system is localized in a small element of approximately 1600 A diameter. The centriole is tentatively proposed as being this control element.     

Structure and Function of Mitochondria in Crithidia fasciculata*

SYNOPSIS. In correlating mitochondrial structure with composition and function of the electron-transport system in Crithidia fasciculata, failure to find cytochrome oxidase in isolated mitochondria coincided with the presence of longitudinally-oriented lamellar cristae in the mitochondria in intact cells. Cytochromes b and c were detected spectrophotometrically. Respiration of intact cells and mitochondria, measured polarographically, was sensitive to 10⁻⁴ M antimycin A and 5 × 10⁻⁴ M KCN. The difficulty in detecting cytochrome oxidase and the biogenesis of mitochondria are briefly discussed.     

Gene mutation eliminating antimycin A-tolerant electron transport in Ustilago maydis

A class of mutants of Ustilago maydis selected on a fungitoxic oxathiin lack of antimycin A-tolerant respiratory system which is present in wild-type cells. This system provides, directly or indirectly, for considerable resistance to antimycin A because growth of mutant cells lacking the system is much more sensitive to the antibiotic than that of the wild type. Antimycin A-sensitive O(2) uptake and growth is found in half of the progeny from crosses of mutant to wild type. All antimycin A-sensitive segregants are somewhat more resistant to oxathiins than the antimycin A-resistant segregants. The respiration of the mutant is strongly inhibited by cyanide and azide at concentrations which stimulate respiration of the wild type. Respiration of both mutant and wild type is about equally inhibited by rotenone. It appears that the mutation alters some component of the respiratory system located between the rotenone inhibition site and the antimycin A inhibition site that permits shift of electron transport to an alternate terminal oxidase when the normal electron transport pathway is blocked.     

Interaction of Colicins with Bacterial Cells IV. Immunity Breakdown Studied with Colicins Ia and Ib

Colicinogenic cells are immune to the lethal effect of the colicin which they produce. In the presence of very high concentrations of colicin, however, colicinogenic cells are no longer immune to the homologous colicin. This phenomenon, immunity breakdown, was studied with colicins Ia and Ib. The biochemical effects of colicin Ib on Escherichia coli were studied with a standard noncolicinogenic strain. At multiplicities of about 10 or higher, colicin Ib inhibited incorporation of leucine into protein and incorporation of (32)P-inorganic phosphate into deoxyribonucleic acid and ribonucleic acid by more than 95%. Under the same conditions, (32)P incorporation into phospholipid and nucleotide fractions was inhibited only partially (about 80 and 60%, respectively). Inhibition of (32)P incorporation into the terminal phosphorus of adenosine triphosphate was also considerably less than that of macromolecular synthesis (50 to 60%). (32)P incorporation into the nonnucleotide organic phosphate fraction was not inhibited. Respiration was not affected. Colicin Ia showed the same biochemical effects as colicin Ib. A mutant of an Ib-colicinogenic E. coli strain selected for resistance to low concentrations of colicin Ia was shown to be resistant to high concentrations of homologous colicin Ib, whereas the parent Ib-colicinogenic strain is sensitive to high concentrations of colicin Ib. This mutant lost its specific receptors for colicin Ib. Moreover, the biochemical effects of high concentrations of colicin Ib on Ib-colicinogenic cells during immunity breakdown were similar to the effects found in sensitive cells exposed to low concentrations of the same colicin. It is concluded that the killing of colicinogenic cells in the presence of high concentrations of homologous colicin is indeed caused by the homologous colicin molecules.     

Respiratory metabolism of normal and divisionless strains of Candida albicans

Respiration of a normal strain of Candida albicans was compared with that of a divisionless mutant which has a biochemical lesion such that metabolically generated hydrogen "spills over," during growth, for non-specific dye reduction. This waste is not at expense of growth, since both strains grow at essentially similar rates, nor at expense of respiration, since the mutant reduces oxygen more rapidly than the normal strain. Respiration in both strains is qualitatively similar, and seemingly unique among highly aerobic organisms in that it is not mediated by cytochrome oxidase. In resting cells of both strains, respiration is not only resistant to, but markedly stimulated by, high concentrations of cyanide, carbon monoxide, and azide. In contrast, growth of these yeasts is inhibited by low concentrations of cyanide and azide. Cytochrome oxidase could not be detected in cell-free preparations; reduced cytochrome c was not oxidized by such preparations. Cytochrome bands could not be observed in thick cell suspensions treated with reducing agents. However, incorporation of superoptimal levels of zinc and iron into the culture medium resulted in growth of cells possessing distinct cytochrome bands; respiration of these cells remained insensitive to cyanide, monoxide, and azide, and the bands were maintained in a reduced form on oxygenation. In the divisionless yeast, tetrazolium dyes compete with oxygen for reduction; this is not the case in the normal strain. The firmness with which hydrogen transfer is channeled in the latter for reduction of disulfide bonds (of importance in the division mechanism) and of oxygen, is contrasted with the lack of such control in the mutant.     

Carbohydrate metabolism of malarial parasites--I. Metabolism of lactate in Plasmodium knowlesi infected monkey erythrocytes

Lactate metabolism by Plasmodium knowlesi infected erythrocytes was examined after careful removal of leucocytes from cell preparations. Infected cells were able to metabolize glucose, pyruvate and lactate. Respiration of infected erythrocytes was maximally stimulated by lactate and to a lesser degree by pyruvate and glucose. Respiration of infected cells was insensitive to stimulation by succinate or glutamate or inhibition by malonate. Mepacrine was found to be a potent respiratory inhibitor. Chromatographic analysis of end products of lactate metabolism showed incorporation of carbon from [2-C14]lactate into phosphoenol pyruvate, 3-phosphoglycerate and malate. Experimental data failed to provide evidence for the presence of a functional citric acid cycle activity in infected cells.     

Das rhythmische Verhalten einer farblosen Mutante von Euglena gracilis

Summary A colorless mutant of Euglena gracilis shows a cireadian rhythmic mobility in darkness just as mixotrophic and autotrophic green forms do.Therefore the obligatory heterotrophic form is very suitable for testing to what extent the rhythmic behavior of the mixotrophic cells, in contrast to that of the autotrophic cells, is independent of photosynthesis.Like the green cells, the colorless cells are synchronized by a single transient from light to dark. About 12 hours after the beginning of the darkness the rhythmic mobility of the colorless cells attains the first maximum, whereas the green forms both show their first maximum 18 hours after the end of the light period, which is much more advantageous for a photosynthesizing organism.The free running period seems to be dependent on the temperature during anaerobic glycolysis and independent of temperature during respiration, just as it has been recently found out in the case of green forms. The type of energy supply changes with the age of the cultures.Respiration has no significance as energy source for the rhythm which continues under pure glycolytic conditions.A sudden increase of the constant temperature does not shift the phase. The same is true in the case of mixotrophic cells but not in the case of autotrophic cells. However, the rhythm is often first suppressed for several days.A lowering of the temperature is followed by two or three transients with about half the frequency, but it does not influence the phase as far as it can be extrapolated. The same has been shown to be true in the case of mixotrophic cells.The rhythmic behavior of the heterotrophic cells is very similar to that of the mixotrophic ones. However, the heterotrophic cells are very sensitive to changes in temperature, which are more compensated for in the mixotrophic cells, apparently by photosynthesis.     

Effect of chilling temperatures on the activities of glyoxysomal and mitochondrial enzymes from castor bean seedlings

Castor bean (Ricinus communis L.) is included among the group of plants sensitive to chilling temperatures. Seedlings of this species were shown to exhibit visible symptoms of this injury, as well as impaired radicle growth and storage product mobilization. Respiration of intact seedlings and oxidation of succinate by isolated mitochondria displayed discontinuities in Arrhenius plots of their reaction velocities, characteristic of chilling species. However, gluconeogenic glyoxysomal enzymes do not display such discontinuities, indicating that there is probably no functional relationship between these enzymes and the glyoxysomal membrane.     

Acetaminophen toxicity results in site-specific mitochondrial damage in isolated mouse hepatocytes

Exposure of isolated mouse hepatocytes to a toxic concentration of acetaminophen (5 mM) resulted in damage to the mitochondrial respiratory apparatus. The nature of this damage was investigated by measuring respiration stimulated by site-specific substrates in digitonin-permeabilized hepatocytes after acetaminophen exposure. Respiration stimulated by succinate at energy-coupling site 2 was most sensitive to inhibition and was decreased by 47% after 1 h. Respiration supported by NADH-linked substrates (site 1) was also decreased but to a lesser extent, while there was no decrease in the rate of ascorbate + N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD)-supported respiration (site 3). The loss of mitochondrial respiratory function was accompanied by a decrease in ATP levels and ATP/ADP ratios in the cytosolic compartment and was preceded by a loss of reduced glutathione in both the cytosol and mitochondria. All these effects occurred well before the loss of cell membrane integrity. The putative toxic metabolite of acetaminophen, N-acetyl-p-benzoquinonimine (NAPQI), produced a similar pattern of respiratory dysfunction in isolated hepatic mitochondria. Respiration stimulated by succinate- and NADH-linked substrates was very sensitive to 50 microM NAPQI, while ascorbate + TMPD-supported respiration was unaffected. The interaction between NAPQI and the respiratory chain was further investigated using submitochondrial particles. Succinate dehydrogenase (associated with respiratory complex II) was found to be very sensitive to NAPQI, while NADH dehydrogenase (respiratory complex I) was inhibited to a lesser extent. Our results indicate that a loss of the ability to utilize succinate- and NADH-linked substrates due to attack of the respiratory chain by NAPQI causes a disruption of energy homeostasis in acetaminophen hepatotoxicity.     

Dexamethasone treatment specifically increases the basal proton conductance of rat liver mitochondria

We investigated the role that mitochondrial proton leak may play in the glucocorticoid-induced hypermetabolic state. Sprague-Dawley rats were injected with dexamethasone over a period of 5 days. Liver mitochondria and gastrocnemius subsarcolemmal and intermyofibrillar mitochondria were isolated from dexamethasone-treated, pair-fed and control rats. Respiration and membrane potential were measured simultaneously using electrodes sensitive to oxygen and to the potential-dependent probe triphenylmethylphosphonium, respectively. Five days of dexamethasone injection resulted in a marked increase in the basal proton conductance of liver mitochondria, but not in the muscle mitochondrial populations. This effect would have a modest impact on energy expenditure in rats.     

Effects of short chain fatty acids on radicle emergence and root growth in lettuce

Abstract. Short chain fatty acids inhibit both radicle emergence and root growth in lettuce. The transition from ineffectual to inhibitory levels occurs abruptly. Root growth is more sensitive to lower concentrations than radicle emergence and is invariant with chain length. The effect of short chain alcohols on radicle emergence is similar to that of short chain acids, but their comparatively severe inhibition of root growth varies with chain length. Alkanes of the same chain lengths have no noticeable effect. Respiration is not altered by a representative short chain fatty acid (heptanoic). Lettuce seeds are sensitized to phytochrome-absorbed light by short chain fatty acids as found by Berrie and co-workers.     

Induction of cytochrome formation and stimulation of oxidative dissimilation by hemin inStreptococcus lactis andLeuconostoc mesenteroides

Aerobic glucose dissimilation of washed cells ofStreptococcus lactis grown in peptone-glucose-yeast extract medium is characterized by the formation of large amounts of lactic acid, a small amount of acetic acid, and traces of acetoin: a corresponding amount of oxygen is taken up. Aerobic metabolism by washed cells ofS. lactis andLeuconostoc mesenteroides is far more oxidative when the cells have been grown on peptone-glucose-yeast extract agar supplied with 10 ppm of hemin than when they have been grown in the absence of hemin. In the former case respiration is strongly inhibited by KCN and only slightly by bis(tributylgermanium) oxide, (Bu₃Ge)₂O. Respiration of cells grown without hemin, on the other hand, is strongly inhibited by (Bu₃Ge)₂O but only moderately by KCN. In cells grown in the presence of hemin, spectra of ana ₂- andb-type cytochrome were recognized but not in cells grown without hemin. The NADH-oxidase activity of such cells is not affected by KCN.Our results strongly suggest that by growth in the presence of hemin a cytochrome-mediated respiration system is induced which replaces, in part, the NADH-oxidase-mediated respiration. Whereas the latter is sensitive to (Bu₃Ge)₂O, the former apparently is little or not. However it is quite sensitive to KCN.When hemin is added to washed cells ofS. lactis grown without hemin the rate of oxygen uptake increases immediately though no cytochromes are present and respiration remains sensitive to (Bu₃Ge)₂O. Possibly hemin stimulates the NADH-oxidase activity of these cells.The author gratefully acknowledges the skilful technical assistance of Mrs. A. J. M. Dekkers-van der Mark and the able performance of gas chromatographic determinations by Miss E. Ch. Th. Gevers and Mrs. G. G. Versluisde Haan.